Reactive oxygen species participate in the control of mouse embryonic cell death.
Programmed cell death or apoptosis is an essential process during the morphogenesis of a large number of structures. Evidence obtained over the past few years indicates that, in some cases, the generation of reactive oxygen species (ROS) is an important event during the course of apoptosis. Using an in vitro culture system in which digit individualization of developing limbs normally occurs, we assayed the effect of different antioxidants on the cell death that takes place at interdigits. The addition of phenol, dimethyl sulfoxide, or 2',7'-dichlorodihydrofluorescein diacetate (DCDHF-DA) to murine developing limbs in culture prevented digit individualization as well as the typical interdigital cell death. Two ROS-sensitive dyes, 3-(4,5-dimethylthiazol)-2,5-diphenyl tetrazolium bromide and DCDHF-DA, stained interdigits and the so-called "necrotic zones," implying that they contain cells under oxidative stress. Very few interdigital cells were doubly stained with the ROS probes and two cell death indicators (i.e., acridine orange and propidium iodide), suggesting that they detect a different stage during the course of apoptosis. Furthermore, we found cells stained for ROS that did not express a specific macrophage marker and in a few cases were seen surrounded by a macrophage. Surprisingly, many regions of the midgestation mouse embryo that are undergoing cell death correlated with those that have a markedly higher level of ROS. Our data suggest that the generation of oxidative stress is a common requirement for cell death that occurs during mouse embryonic development.
Salas-Vidal
E
E
Departmento de Genética y Fisiología Molecular, Universidad Nacional Autónoma de México, Cuernavaca, Morelos, México.
Lomelí
H
H
Castro-Obregón
S
S
Cuervo
R
R
Escalante-Alcalde
D
D
Covarrubias
L
L
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004622:Embryo, Mammalian; D005314:Embryonic and Fetal Development / Q000187:drug effects / Q000502:physiology; D005452:Fluoresceins / Q000494:pharmacology*; D005456:Fluorescent Dyes; D018878:Limb Buds / Q000166:cytology* / Q000502:physiology* / Q000648:ultrastructure; D008264:Macrophages / Q000166:cytology / Q000502:physiology / Q000648:ultrastructure; D051379:Mice; D018613:Microscopy, Confocal; D009024:Morphogenesis / Q000187:drug effects; D009924:Organ Culture Techniques; D019800:Phenol / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000502:physiology*
C110400:2',7'-dichlorodihydrofluorescein diacetate; D000975:Antioxidants; D005452:Fluoresceins; D005456:Fluorescent Dyes; D017382:Reactive Oxygen Species; D019800:Phenol; D004121:Dimethyl Sulfoxide